The majority of excitatory synapses in the mammalian brain are located on dendritic spines. Changes in spine structure and function play critical roles in brain development and plasticity. Conversely, alterations in spine morphogenesis occur in psychiatric disorders, including schizophrenia. Consequently, understanding the molecular mechanisms underlying spine plasticity and their alterations in psychiatric disorders could lead to the rational development of novel therapeutic strategies in mental disorders. Kalirin, a neuronal guanine-nucleotide exchange factor (GEF) for small GTPases, is emerging as a regulatory hub of structural and functional plasticity in spines. Recent genetic and neuropathological studies in human subjects have also implicated kalirin and its signaling partners in psychiatric disorders, including schizophrenia. Recent evidence has shown that kalirin isoforms are differentially affected in the brains of human subjects with mental disorders, carry missense mutations enriched in mental disorders, and have differential impacts on dendrite and spine morphology. Moreover, kalirin isoforms interact with several important trafficking proteins. However, the roles of these isoforms and their interactions in spines have not been examined. Based on these findings, we hypothesize that kalirin isoforms are distributed in distinct postsynaptic microcompartments, where they play specific functions in coordinating synaptic glutamate receptor trafficking with spine morphogenesis. Recent developments in superresolution imaging have opened up novel directions for functional analysis of synaptic proteins. Here we will utilize superresolution imaging in combination with molecular manipulations, biochemistry, immune-electron microscopy, and electrophysiology to pursue the following aims: 1) To map the nanoscale organization of kalirin isoforms in postsynaptic microcompartments. 2) To characterize the molecular mechanisms underlying the interactions of kalirin isoforms with Arf6, SNX1/2 and dynamin. 3) To determine the consequences of the interaction of kalirin isoforms with Arf6, SNX1/2, and dynamin on postsynaptic structure and function. The proposed studies will be the first to investigate at nanoscopic resolution the sub-synaptic distribution and functions of isoforms of an important regulator of spine plasticity and pathology, providing insight into the roles of protein isoforms i synapses. This proposal will also investigate novel roles for kalirin isoforms in coordinating synaptic trafficking with spine remodeling, which could implicate altered synaptic trafficking as a candidate pathophysiological mechanism in some mental disorders. The proposal will therefore shed light on key mechanisms of synaptic plasticity and pathology, and can facilitate the rational development of novel therapeutic strategies aimed at reversing synaptic deficits in mental disorders.